Abstract

Background

High abundance protein depletion is a major challenge in the study of serum/plasma
proteomics. Prior to this study, most commercially available kits for depletion of
highly abundant proteins had only been tested and evaluated in adult serum/plasma,
while the depletion efficiency on umbilical cord serum/plasma had not been clarified.
Structural differences between some adult and fetal proteins (such as albumin) make
it likely that depletion approaches for adult and umbilical cord serum/plasma will
be variable. Therefore, the primary purposes of the present study are to investigate
the efficiencies of several commonly-used commercial kits during high abundance protein
depletion from umbilical cord serum and to determine which kit yields the most effective
and reproducible results for further proteomics research on umbilical cord serum.

Results

The immunoaffinity based kits (PROTIA-Sigma and 5185-Agilent) displayed higher depletion
efficiency than the immobilized dye based kit (PROTBA-Sigma) in umbilical cord serum
samples. Both the PROTIA-Sigma and 5185-Agilent kit maintained high depletion efficiency
when used three consecutive times. Depletion by the PROTIA-Sigma Kit improved 2DE
gel quality by reducing smeared bands produced by the presence of high abundance proteins
and increasing the intensity of other protein spots. During image analysis using the
identical detection parameters, 411 ± 18 spots were detected in crude serum gels,
while 757 ± 43 spots were detected in depleted serum gels. Eight spots unique to depleted
serum gels were identified by MALDI- TOF/TOF MS, seven of which were low abundance
proteins.

Conclusions

The immunoaffinity based kits exceeded the immobilized dye based kit in high abundance
protein depletion of umbilical cord serum samples and dramatically improved 2DE gel
quality for detection of trace biomarkers.